Opioids have been used to relieve pain for centuries. However, the abuse of opioids is a major health care problem in the world. The bases for the behavioral effects of these drugs of abuse will be explained by studies of their effects on brain systems, and then ultimately, on specific ion or neurotransmitter receptor channels. It is the long term goal of the proposed project to focus on the latter aspect, to determine the action of opioids on ion and neurotransmitter receptor channels. On primary afferent neurons, kappa and mu opioid receptors are coupled to specific voltage-dependent calcium channels which likely regulate release of neurotransmitter from primary afferent terminals. We are interested in the coupling of kappa and mu opioid receptors to specific calcium channels and the regulation of those calcium channels by phosphorylation. Our specific aims are to determine: 1) which pertussis toxin sensitive G protein couples kappa receptors to calcium channels using specific antibodies directed at the G protein alpha subunits, alphao, alphai1, alphai2 and alphai3, to block the coupling; 2) the specific high threshold calcium channel coupled to mu and kappa receptors using a combination of specific calcium channel blockers and voltage clamp protocols; 3) which high threshold calcium currents are regulated by phosphorylation by protein kinase A (PKA) or protein kinase C (PKC); and 4) whether phosphorylation by PKA and PKC regulates the coupling between opioid receptors and calcium channels. The hypotheses to be tested are that mu and kappa receptors couple to a specific high threshold transient calcium channel by a specific G protein and that phosphorylation of the calcium channel increases the number of channels available for regulation and enhances the opioid receptor reduction of calcium current.